Bacterialpathogenscanpersistoverthelifetimeoftheirhostinagrowthrestricted,dormantstate.Pathogens likeClostridiumdifficile,thatcansurviveassporesareespeciallyresistanttoantibiotics.C.difficileisestimated tocosttheUShealthcaresystemmorethanonebilliondollarsperyear.Understandinghowbacteriaregulate dormancyisessentialtoourabilitytotreatthesekindsofinfection.Tobecomequiescent,cellsmustshutdown proteinsynthesis.Post-translationalmodificationsoffactorsinvolvedintranslationarelikelytoplayarolein thisregulation.TheproposedworkaimstodeterminetheroleofelongationfactorG(EF-G)phosphorylationin regulatingribosomerecyclingduringdormancy.EF-Gisstablyphosphorylatedinregionsthatinteractwith ribosomerecyclingfactor(RRF)duringstationaryphaseinmanybacteria.EF-Gisalsoknowntobestably phosphorylatedinthemodelspore-formerBacillussubtilis.Usingbiochemicalapproaches,wewillinvestigate theeffectsoftheseconservedmodificationsonribosomerecycling,andontheGTPaseactivityofEF-G.The roleofEF-Gintherecyclingofribosomessequesteredbyhibernationfactorswillalsobedetermined.Using singlemoleculefluorescenceenergytransfer(smFRET)approaches,wewillinvestigatetheshort-lived interactionsofP~EF-GwithRRFonindividualribosomesthathaveterminatedtranslation.Finally,sincelittleis knownabouthowtranslationisinhibitedinsporeswewillprofiletheassociationofribosomalsubunitsandtheir activityoverthecourseofsporedevelopmentandasthesporegerminates.Wewillalsoidentifyphosphosites onEF-Ginthesporeanddeterminetheeffectsofthesemodificationsontherecyclingandelongationactivity ofEF-Ginvitro.Thesestudieswillimpactourunderstandingofribosomerecyclingandhowphosphorylation canbeusedtoregulatedormancy.

Public Health Relevance

Manypathogensthatareathreattohumanhealthuseagrowth-restricted,dormant statefortransmission,persistence,pathogenesis,andantibioticevasion.Thisworkwill investigatehowproteinsynthesis,andribosomerecyclinginparticular,canbeusedto regulatebacterialdormancy.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32GM122266-01
Application #
9258129
Study Section
Special Emphasis Panel (ZRG1-F13-C (20)L)
Program Officer
Melillo, Amanda A
Project Start
2017-02-01
Project End
2020-01-31
Budget Start
2017-02-01
Budget End
2018-01-31
Support Year
1
Fiscal Year
2017
Total Cost
$56,334
Indirect Cost
Name
Columbia University (N.Y.)
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032